DESCRIPTION: HIV transmission between intravenous drug users is caused by the sharing of needles contaminated with HIV-infected cells or cell-free virus. The pathway that HIV takes following intravenous injection has been presumed to lead immediately to widespread dissemination, bypassing mucosal barrier defenses. By contrast, we have observed a strong initial filtering of HIV-infected cells in the lungs of humanized mice challenged intravenously wit HIV. The effect resembles T cell trapping that is observed in immunotherapy studies in humans and in T cell adoptive transfer experiments in mice, where injected activated lymphocytes first accumulate in the lung. The lung is already known to act as a filter against endogenous or exogenous venous emboli, preventing them from damaging peripheral tissues when injected into the venous circulation. In this proposal, we explore our preliminary findings that indicate cell-associated HIV localizes initially to the lung in the humanized mouse. Recent humanized mouse studies have observed that spread from organ to organ is mediated by infected cells that may use cell-to-cell contacts to initiate new infections rather than by cell-free virus. We will examine the migration of HIV-infected cells into the lung and follow the phenotypic and functional changes that accompany spread to distal sites during parenteral transmission. To study acute intravenous transmission, we have developed novel tools to examine the role of cell migration and cell-cell interactions in HIV dissemination. We are studying the intravenous inoculation of humanized mice, which carry a human immune system with CD4+ T cells that support HIV infection. We have designed novel recombinant fluorescent HIV clones with Env from transmitted founder viruses, that are more robustly replication-competent than lab isolates and allow us to track infected cells in the initial minutes to hours after intravenous infection. Fluorescent viral clones allow us to measure infection with flow cytometry and to examine anatomical compartmentalization and formation of cell-cell infectious structures called virological synapses (VS) in tissues using two-photon live microscopy. We will examine whether prolonged trapping in the lung can inhibit intraveneous transmission. We examine the pathway of cell-free and cell-associated inoculums that are introduced through the intraveneous route. Antibodies that preferentially inhibit cell-free transmission will help to assess the contribution of cell-cel infection in parenteral transmission. We will study what makes a transmitted founder R5-tropic virus infect more efficiently through intravenous routes than an isogenic lab- adapted X4-tropic clone. We examine the overarching hypothesis that the lung is a major anatomical barrier that plays a crucial role in the establishment of intravenous infection and that both cell migration and cell-to-cell infection through VS are critical for the establishment of these infections.